Human pluripotent stem cells are promising and potentially unlimited cell source for regenerative medicine; however, tumor formation is of major concern for cell therapy with pluripotent stem cells. Residual undifferentiated stem cells in preparations of stem cell derivatives, including cardiomyocyte differentiation cultures, have potential to proliferate and form teratoma upon transplantation, posing a major hurdle for safe cell therapy. Careful assessment of differentiation cultures and development of strategies to remove undifferentiated stem cells will facilitate qualification of final stem cell products for therapeutic use. In this study, we propose to develop sensitive, nanotechnology-based assays to detect residual undifferentiated cells, characterize cardiomyocyte differentiation cultures at various stages for their propensities to form teratomas, and evaluate strategies to selectively remove residual undifferentiated cells. Technology established in this study will be valuable for the development of novel cell therapy using human pluripotent stem cells.